Gene therapy of bladder pain with herpes simplex virus (HSV) vectors expressing preproenkephalin (PPE).
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Afferent Pathways↗
Biomedical subjects
W C de Groat
Publications and source records attributed to W C de Groat.
Suppression of the tetrodotoxin-resistant sodium channel (PN3/SNS): a possible new treatment for bladder pain.
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Acetic Acid↗
Protein kinase C is involved in M1-muscarinic receptor-mediated facilitation of L-type Ca2+ channels in neurons of the major pelvic ganglion of the adult male rat.
We used patch clamp recording techniques to determine if muscarinic signaling mechanisms are present in dissociated autonomic neurons obtained from the major pelvic ganglion, which provides the cholinergic innervation of the urinary bladder and other pelvic organs. The M1 specific agonist, McN-A-343 (2-30 microM) enhanced Ca2+ currents in approximately 37% of neurons (by 50-80%). This enhancement was reduced by atropine (5-10 microM) or a PKC inhibitor (bisindolylmaleimide, 50-200 nM). In responsive neurons Ca2+ currents were also enhanced by the phorbol ester, phorbol-12,13-dibutyrate (50-300 nM) and the dihydropyridine agonist Bay K 8644 (5 microM) and had kinetics of activation and inactivation as expected for L-type Ca2+ channels. We conclude that in a subpopulation of MPG neurons, M1-mediated activation of PKC phosphorylates and enhances L-type Ca2+ channel activities. This muscarinic facilitatory mechanism in MPG neurons may be the same as the M1-mediated facilitation of transmitter release reported previously at the nerve terminals in the urinary bladder.
Animals↗
Neural control of the urethra.
Coordination between the urinary bladder and the urethra is mediated by multiple reflex pathways organized in the brain and spinal cord. Some reflexes promote urine storage; whereas other reflexes facilitate voiding. During bladder filling, activation of mechanoreceptor afferent nerves in the bladder wall triggers firing in the cholinergic efferent pathways to the external urethral sphincter (EUS) and in sympathetic adrenergic pathways to the urethral smooth muscle. These storage reflexes are dependent upon interneuronal circuitry in the spinal cord. During voiding the spinal storage reflexes are inhibited by supraspinal mechanisms which originate in the pontine micturition center. Glutamatergic, serotonergic and alpha, adrenergic excitatory transmission as well as GABAergic/glycinergic inhibitory transmission have been implicated in the central control of sphincter reflexes. During voiding, a parasympathetic nitrergic inhibitory input to the urethral smooth is activated. This reflex mechanism which is triggered by bladder afferents persists in paraplegic rats and therefore must be mediated at least in part by spinal interneuronal circuitry. In female rats, the parasympathetic nitrergic pathway is prominent; but in male rats it is obscured by a dominant parasympathetic cholinergic excitatory input to the urethral smooth muscle. The function of the cholinergic pathway in voiding is uncertain. Stimulation of urethral afferents can also influence bladder activity. Contraction of the external urethral sphincter activates afferents that inhibit reflex bladder contractions; whereas infusion of fluid through the urethra facilitates bladder contractions. These reflexes are also organized in the spinal cord and presumably play a role in urine storage and elimination. Alterations in primitive bladder-to-urethra and urethra-to-bladder reflex mechanisms may contribute to neurogenic bladder dysfunction.
Afferent Pathways↗
Autologous primary muscle-derived cells transfer into the lower urinary tract.
The goal of these experiments was to establish the basic methodology for future clinical applications of muscle-derived cells (MDC) tissue engineering and gene transfer for the treatment of urological dysfunction. Primary MDC isolated via preplating techniques from adult female SD rats were transduced with retrovirus encoding the expression of beta-galactosidase reporter gene. The MDC were injected into the right and left lateral walls of the bladder and proximal urethra of the autologous animals (n = 6) with a 10 microl Hamilton micro syringe. The amount of injected MDC ranged from 1 to 2 x 10(6) cells. The injected tissue was harvested after 7, 14, and 28 days, sectioned and examined histologically for beta-galactosidase and immunohistochemically for fast myosin heavy chain specific to skeletal muscle. The tissues were also stained for anti-CD4 and anti-CD8 antibodies to assess for cellular immune reaction. We have detected a large number of autologous MDC expressing beta-galactosidase and positively stained for fast myosin heavy chain in the bladder and urethral wall. Many injected myoblasts and myotubes were also seen in the bladder and urethral wall at each time point. Staining of lymphocytes with anti-CD4 and anti-CD8 antibodies was negative after MDC injection at each time point. We have demonstrated the long-term survival of autologous MDC and MDC mediated gene transfer into the bladder and urethral wall. Autologous MDC and MDC mediated gene transfer may be a promising treatment to augment bladder and urethral sphincter function.
Animals↗
Nitric oxide synthase gene therapy for erectile dysfunction: comparison of plasmid, adenovirus, and adenovirus-transduced myoblast vectors.
BACKGROUND AND PURPOSE: Nitric oxide (NO) has been recognized as an important transmitter for genitourinary tract function. This transmitter mediates smooth muscle relaxation and is essential for erection. The objective of our research was to determine whether overexpression of nitric oxide synthase (NOS) in the corpus cavernosum of the penis would correct erectile dysfunction. MATERIALS AND METHODS: We introduced the inducible form of the enzyme NOS (iNOS) into the corpus cavernosum of adult (250-300 g) male Sprague-Dawley rats by injecting a solution of plasmid, adenovirus, or adenovirus-transduced myoblast cells (adeno-myoblast) (N = 3-5 each group). We also injected plasmid, adenovirus, and adeno-myoblast encoding the expression of the beta-gatactosidase reporter gene. RESULTS: We noted expression of beta-galactosidase throughout the corpora cavernosum after injection of each of the three solutions. Staining was greatest for adeno-myoblast followed by adenovirus and then plasmid. The basal intracavernous pressure (ICP) of iNOS-treated animals (adenovirus and adenovirus-transduced myoblast) increased to 55 +/- 23 cm H(2)O v 5 +/- 6 H(2)O in naive animals (P = 0.001). Stimulation of the cavernous nerve (15 Hz, 1.5 msec, 10-40 V, 1 min) resulted in a twofold increase in ICP (adenovirus and adeno-myoblast) from the basal level of the iNOS-treated animals. Direct in situ measurement of NO demonstrated release of 1 to 1.3 microM NO in the adeno-myoblast-treated penis. CONCLUSION: Myoblast-mediated gene therapy was more successful in delivering iNOS into the corpus cavernosum than were the direct adenovirus or plasmid transfection methods. Gene therapy of NOS may open new avenues of treatment for erectile dysfunction. Control of NOS expression would be necessary to prevent priapism.
Adenoviridae↗
Persistence and survival of autologous muscle derived cells versus bovine collagen as potential treatment of stress urinary incontinence.
PURPOSE: We explored the use of autologous muscle derived cells as a method of treating stress urinary incontinence. We determined whether urethral muscle derived cell injection is feasible and compared it with bovine collagen injection. MATERIALS AND METHODS: Muscle derived cells isolated from female Sprague-Dawley rats were first transduced with retrovirus carrying the transgene for beta-galactosidase. We injected approximately 1 to 1.5 x 106 cells into the bladder wall and proximal urethra of 6 autologous animals. Tissue was harvested after 3 and 30 days, sectioned, stained for fast myosin heavy chain and assayed for beta-galactosidase. To compare muscle derived cell and bovine collagen injections 100 microl. of commercially available bovine collagen were also injected in Sprague-Dawley female rats. Tissue was harvested in 3 animals each after 3 and 30 days, sectioned and stained for trichrome. Subsequently, 3 adult SCID mice were used to compare the level of transgene expression at each time point after injecting 1.5 x 106 cells per injection, which were transduced with adenovirus carrying the transgene for beta-galactosidase. RESULTS: A large number of cells expressing beta-galactosidase were observed in the bladder and urethral wall 3 and 30 days after autologous cell injection in Sprague-Dawley rats. The persistence of primary muscle derived cells at 3 days was similar to that of collagen. However, at 30 days there was significant cell persistence while only a minimal amount of injected bovine collagen was detectable. Approximately 88% of the beta-galactosidase expression at day 3 remained at day 30 in SCID mice. CONCLUSIONS: We present 2 new findings important for the emerging field of urological tissue engineering, including the feasibility of injecting autologous skeletal muscle derived cells into the lower urinary tract and the greater persistence of such injected cells versus injected bovine collagen. Therefore, autologous muscle derived cell injection may be an attractive alternative treatment option for stress urinary incontinence.
Animals↗
Pharmacology of the lower urinary tract.
The functions of the lower urinary tract, to store and periodically release urine, are dependent on the activity of smooth and striated muscles in the urinary bladder, urethra, and external urethral sphincter. This activity is in turn controlled by neural circuits in the brain, spinal cord, and peripheral ganglia. Various neurotransmitters, including acetylcholine, norepinephrine, dopamine, serotonin, excitatory and inhibitory amino acids, adenosine triphosphate, nitric oxide, and neuropeptides, have been implicated in the neural regulation of the lower urinary tract. Injuries or diseases of the nervous system, as well as drugs and disorders of the peripheral organs, can produce voiding dysfunctions such as urinary frequency, urgency, and incontinence or inefficient voiding and urinary retention. This chapter will review recent advances in our understanding of the pathophysiology of voiding disorders and the targets for drug therapy.
Animals↗
Analysis of the afferent limb of the vesicovascular reflex using neurotoxins, resiniferatoxin and capsaicin.
The afferent limb of the vesicovascular reflex (VV-R) evoked by distension or contraction of the urinary bladder (UB) was studied in urethane-anesthetized female rats by examining the changes in VV-R after administration of C-fiber afferent neurotoxins [capsaicin and resiniferatoxin (RTX)]. Systemic arterial blood pressure increased parallel (5.1 to 53.7 mmHg) with graded increases in UB pressure (20 to 80 cm H(2)O) or during UB contractions. The arterial pressor response to UB distension was significantly reduced (60-85%) by acute or chronic (4 days earlier) intravesical administration of RTX (100-1,000 nM) or by capsaicin (125 mg/kg sc) pretreatment (4 days earlier). Chronic neurotoxin treatments also increased the volume threshold (>100%) for eliciting micturition in anesthetized rats but did not change voiding pressure. Acute RTX treatment (10-50 nM) did not alter the arterial pressor response during reflex UB contractions, whereas higher concentrations of RTX (100-1,000 nM) blocked reflex bladder contractions. It is concluded that VV-R is triggered primarily by distension- and contraction-sensitive C-fiber afferents located, respectively, near the luminal surface and deeper in the muscle layers of the bladder.
Administration, Intravesical↗
Nitric oxide modulates Ca(2+) channels in dorsal root ganglion neurons innervating rat urinary bladder.
The effect of a nitric oxide (NO) donor on high-voltage-activated Ca(2+) channel currents (I(Ca)) was examined using the whole cell patch-clamp technique in L(6)-S(1) dorsal root ganglion (DRG) neurons innervating the urinary bladder. The neurons were labeled by axonal transport of a fluorescent dye, Fast Blue, injected into the bladder wall. Approximately 70% of bladder afferent neurons exhibited tetrodotoxin (TTX)-resistant action potentials (APs), and 93% of these neurons were sensitive to capsaicin, while the remaining neurons had TTX-sensitive spikes and were insensitive to capsaicin. The peak current density of nimodipine-sensitive L-type Ca(2+) channels activated by depolarizing pulses (0 mV) from a holding potential of -60 mV was greater in bladder afferent neurons with TTX-resistant APs (39.2 pA/pF) than in bladder afferent neurons with TTX-sensitive APs (28.9 pA/pF), while the current density of omega-conotoxin GVIA-sensitive N-type Ca(2+) channels was similar (43-45 pA/pF) in both types of neurons. In both types of neurons, the NO donor, S-nitroso-N-acetylpenicillamine (SNAP) (500 microM), reversibly reduced (23.4-26.6%) the amplitude of I(Ca) elicited by depolarizing pulses to 0 mV from a holding potential of -60 mV. SNAP-induced inhibition of I(Ca) was reduced by 90% in the presence of omega-conotoxin GVIA but was unaffected in the presence of nimodipine, indicating that NO-induced inhibition of I(Ca) is mainly confined to N-type Ca(2+) channels. Exposure of the neurons for 30 min to 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10 microM), an inhibitor of NO-stimulated guanylyl cyclase, prevented the SNAP-induced reduction in I(Ca). Extracellular application of 8-bromo-cGMP (1 mM) mimicked the effects of NO donors by reducing the peak amplitude of I(Ca) (28.6% of reduction). Action potential configuration and firing frequency during depolarizing current pulses were not altered by the application of SNAP (500 microM) in bladder afferent neurons with TTX-resistant and -sensitive APs. These results indicate that NO acting via a cGMP signaling pathway can modulate N-type Ca(2+) channels in DRG neurons innervating the urinary bladder.
8-Bromo Cyclic Adenosine Monophosphate↗
Excitatory synaptic currents in lumbosacral parasympathetic preganglionic neurons elicited from the lateral funiculus.
Excitatory postsynaptic currents (EPSCs) in parasympathetic preganglionic neurons (PGNs) were examined using the whole cell patch-clamp recording technique in L6 and S1 spinal cord slices from neonatal rats (6-16 days old). PGNs were identified by labeling with retrograde axonal transport of a fluorescent dye (Fast Blue) injected into the intraperitoneal space 3-7 days before the experiment. Synaptic responses were evoked in PGNs by field stimulation of the lateral funiculus (LF) in the presence of bicuculline methiodide (10 microM) and strychnine (1 microM). In approximately 40% of the cells (total, 100), single-shock electrical stimulation of the LF elicited short, relatively constant latency [3.0 +/- 0.1 (SE) ms] fast EPSCs consistent with a monosynaptic pathway. The remainder of the cells did not respond to stimulation. At low intensities of stimulation, the EPSCs often occurred in an all-or-none manner, indicating that they were mediated by a single axonal input. Most cells (n = 33) exhibited only fast EPSCs (type 1), but some cells (n = 8) had fast EPSCs with longer, more variable latency polysynaptic EPSCs superimposed on a slow inward current (type 2). Type 1 fast synaptic EPSCs were pharmacologically dissected into two components: a transient component that was blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 5 microM), a non-NMDA glutamatergic antagonist, and a slow decaying component that was blocked by 2-amino-5-phosphonovalerate (APV, 50 microM), a NMDA antagonist. Type 2 polysynaptic currents were reduced by 5 microM CNQX and completely blocked by combined application of 5 microM CNQX and 50 microM APV. The fast monosynaptic component of type 1 EPSCs had a linear current-voltage relationship and reversed at a membrane potential of 5.0 +/- 5.9 mV (n = 5), whereas the slow component exhibited a negative slope conductance at holding potentials greater than -20 mV. The type 1, fast synaptic EPSCs had a time to peak of 1.4 +/- 0.1 ms and exhibited a biexponential decay (time constants, 5.7 +/- 0.6 and 38.8 +/- 4.0 ms). In the majority of PGNs (n = 11 of 15 cells), EPSCs evoked by electrical stimulation of LF exhibited paired-pulse inhibition (range; 25-33% depression) at interstimulus intervals ranging from 50 to 120 ms. These results indicate that PGNs receive monosynaptic and polysynaptic glutamatergic excitatory inputs from axons in the lateral funiculus.
2-Amino-5-phosphonovalerate↗
The role of bladder afferent pathways in bladder hyperactivity induced by the intravesical administration of nerve growth factor.
PURPOSE: Interstitial cystitis, a chronic disease of the bladder, is characterized by urinary frequency, urgency and suprapubic pain. Nerve growth factor is a substance that may sensitize afferent nerves and induce bladder hyperactivity. It is often increased in the urine of patients with interstitial cystitis. We evaluated the role of Adelta and C fiber afferents in the type of bladder hyperactivity induced by the intravesical administration of nerve growth factor. MATERIALS AND METHODS: A total of 22 Wistar and 8 Sprague-Dawley adult female rats were anesthetized with 1.2 gm/kg urethane given subcutaneously. A transurethral catheter was inserted into the bladder. Some animals were pretreated with 125 mg/kg capsaicin injected subcutaneously 4 days before nerve growth factor administration. Cystometry was performed by slowly filling the bladder at a rate of 0.04 ml per minute for 15 minutes with a volume of up to 0.6 ml. Parameters measured included volume threshold and pressure threshold for inducing the micturition reflex, compliance, bladder contraction amplitude, number of contractions and the inter-contraction interval. Nerve growth factor (0.5 ml of 20 microg/ml in 10% dimethyl sulfoxide) or a vehicle solution (0.5 ml of 10% dimethyl sulfoxide) was infused into the bladder through a transurethral catheter and retained for 1 hour. RESULTS: In Wistar rats nerve growth factor increased the mean number of contractions by 111% versus controls (5.7 versus 2.7, p <0.05), and decreased the mean volume threshold by 41% (0.244 versus 0.412 ml, p <0.05). This effect of nerve growth factor was not detected in Sprague-Dawley rats. Capsaicin pretreatment increased the volume threshold by 59% but did not change nerve growth factor induced bladder hyperactivity. CONCLUSIONS: The intravesical application of nerve growth factor acutely induced bladder hyperactivity in Wistar but not in Sprague-Dawley rats. Because the C fiber afferent neurotoxin capsaicin did not change the effect of nerve growth factor, we believe that Adelta afferent neurons have a major role in nerve growth factor induced bladder hyperactivity.
Administration, Intravesical↗
Herpes simplex virus mediated nerve growth factor expression in bladder and afferent neurons: potential treatment for diabetic bladder dysfunction.
PURPOSE: Diabetic cystopathy resulting from sensory neuropathy may potentially be treated by direct gene therapy. It has been suggested that nerve growth factor (NGF) has an ameliorative effect in preventing the death in diabetes of afferent dorsal root ganglion neurons, which control bladder function. We investigated NGF gene transfer to the bladder and bladder afferent pathways for treating diabetic cystopathy. We used replication competent and replication defective herpes simplex virus type 1 (HSV-1) vectors that express a functionally active form of the beta-subunit of mouse NGF (beta-NGF) to examine the level and duration of therapeutic gene expression after administration of the vectors. MATERIALS AND METHODS: NGF expression during acute (3 days) and latent (21 days) infections was assessed by enzyme-linked immunosorbent assay (ELISA) and immunohistochemical testing after the injection of 1 x 106 to 1 x 108 pfu HSV-NGF expression vectors into the bladder wall of adult rats. RESULTS: HSV vectors with the strong human cytomegalovirus immediate early promoter used to drive beta-NGF gene expression exhibited increased NGF 3 days after infection in the bladder and L6 to S1 dorsal root ganglia, where bladder afferent neurons are located. ELISA analysis revealed that NGF in the bladder tissue and dorsal root ganglia was increased 7 to 9 and 2 to 4-fold, respectively, over the control vector. Increased NGF expression in L6 to S1 dorsal root ganglia neurons was also detected by immunohistochemical staining with antiNGF antibodies. Extended NGF expression was detected by ELISA 21 days after injection. Replication defective vectors containing HSV-1 latency promoter (LAP-2) driving NGF expressed NGF in the bladder and dorsal root ganglia 21 days after bladder injection. ELISA analysis confirmed an approximate 2 to 3-fold increase of NGF expression in the bladder and L6 to S1 dorsal root ganglia. CONCLUSIONS: The NGF gene may be transferred and expressed in the bladder and bladder afferent pathways using HSV vectors. To our knowledge our study represents the first demonstration of the effectiveness of gene therapy for altering neurotrophic expression in visceral sensory neurons. This technique of gene transfer may be useful for treating certain types of neurogenic bladder dysfunction, such as diabetic cystopathy, in which decreased NGF transport may be a causative factor.
Animals↗
Central muscarinic mechanisms regulating voiding in rats.
The influence of muscarinic receptor stimulation and blockade on the central regulation of micturition was evaluated in conscious female rats. Saline was infused into the bladder to induce repeated bladder contractions and voiding. Increasing doses of a muscarinic agonist, oxotremorine-M (OXO-M; 0.01 to 1 microg/rat) or antagonist, atropine (0.1 to 30 microg/rat) were administered. Intrathecal OXO-M (0.1 microg) increased bladder capacity (BC; 85 +/- 17%), but did not change maximal voiding pressure (MVP), pressure threshold (PT), postvoiding intravesical pressure, or voiding efficiency (VE). Intracerebroventricular OXO-M (0.1 microg) increased BC (97 +/- 6%), MVP (45 +/- 19%), PT (158 +/- 49%), and reduced VE (-17 +/- 5%). A larger dose of OXO-M (1 microg, either i.c.v. or i.t.) produced greater changes. These effects were not reproduced by i.v. injections of OXO-M. The effects of OXO-M were blocked by pretreatment with atropine in a dose (1 microg i.c.v. or i.t.), which alone had no effect on voiding parameters. A larger dose of atropine (10 microg) reduced MP (-31 +/- 7% i.c.v. and -34 +/- 6% i.t.) and VE (-21 +/- 3% i.c.v. and -25 +/- 5% i.t.) but increased BC (52 +/- 8% i.c.v.). These results indicate that activation of muscarinic receptors in the brain or spinal cord can suppress voluntary voiding, but also stimulates bladder activity during bladder filling. The muscarinic inhibitory mechanisms do not appear to be tonically active. The effects of atropine (i.c.v. and i.t.) indicate that muscarinic excitatory mechanisms are tonically active. These findings raise the possibility that voiding function is regulated by both inhibitory and excitatory cholinergic mechanisms in the central nervous system.
Animals↗
Role of spinal alpha(1)-adrenergic mechanisms in the control of lower urinary tract in the rat.
The role of spinal alpha(1)-adrenergic mechanisms in the control of urinary bladder function was examined in urethane (1.2 g/kg s.c.) anesthetized and decerebrate unanesthetized female Sprague-Dawley rats (250-320 g). Bladder activity was recorded via a transurethral catheter during continuous infusion (0.21 ml/min) cystometrograms or under isovolumetric conditions. All drugs were administered intrathecally at the L(6)-S(1) segmental level of spinal cord. During cystometrograms, 3 or 30 nmol of phenylephrine (alpha(1)-adrenergic agonist) did not alter bladder activity; whereas 300 nmol increased the intercontraction interval by 98% and pressure threshold for inducing micturition by 115%, but did not change bladder contraction amplitude. A large dose of phenylephrine (3000 nmol) completely blocked reflex voiding and induced overflow incontinence at a high baseline pressure (mean: 33 cmH(2)O; range: 28-42 cmH(2)O). Under isovolumetric conditions, 3-30 nmol of phenylephrine abolished bladder activity for 22-45 min; whereas smaller doses (0.003-0.3 nmol) were inactive. Doxazosin (50 nmol), an alpha(1)-adrenergic antagonist, decreased intercontraction intervals but did not change bladder contraction amplitude during cystometrograms. Under isovolumetric conditions this dose of doxazosin increased bladder contraction frequency and decreased bladder contraction amplitude. Smaller doses (5 or 25 nmol) of doxazosin did not alter bladder activity. These studies suggest that two types of spinal alpha(1)-adrenergic mechanisms are involved in reflex bladder activity: (1) inhibitory control of the frequency of voiding reflexes presumably by regulating afferent processing in the spinal cord and (2) facilitatory modulation of the descending limb of the micturition reflex pathway.
Adrenergic alpha-Agonists↗
Electrophysiological properties of lumbosacral preganglionic neurons in the neonatal rat spinal cord.
The electrophysiological properties of parasympathetic preganglionic neurons (PGN) in L6 and S1 spinal cord slices from neonatal rats were studied using the patch clamp techniques. PGN were identified by retrograde axonal transport of a fluorescent dye (Fast Blue) injected intraperitoneally before the experiment. PGN in the intermediolateral region of the spinal cord were divided into two classes (tonic PGN and phasic PGN) on the basis of firing properties during prolonged (300 ms) depolarizing current pulses. Tonic neurons exhibited a prolonged discharge (average maximum: 5.6); whereas phasic PGN fired on average only 1.4 spikes during depolarizing pulses. PGN were usually oval in shape. The mean long axis of tonic PGN (20.7+/-0.5 microm) was significantly (P<0.05) larger than that of phasic PGN (16.7+/-0.3 microm). Tonic and phasic PGN had similar resting membrane potentials, thresholds for spike activation, input resistances and action potential durations. The duration of the after-hyperpolarization (AHP) in tonic PGN (200.5+/-11.9 ms) was longer than in phasic PGN (137.6+/-9.8 ms). 4-aminopyridine (4-AP, 0. 5 mM) reduced the threshold for spike activation in tonic and phasic PGN. 4-AP also unmasked tonic firing in phasic PGN (average maximum: 5.5 spikes during 300 ms depolarizing current pulses) and increased firing frequency by 19% in tonic PGN. These data indicate that the different discharge patterns of parasympathetic PGN are dependent in part on differences in the expression of 4-AP-sensitive K(+) channels. The two types of PGN may provide an innervation to different targets in the pelvic viscera.
4-Aminopyridine↗
Preliminary results of myoblast injection into the urethra and bladder wall: a possible method for the treatment of stress urinary incontinence and impaired detrusor contractility.
The purpose of this study is to explore the feasibility of myoblasts, the precursors of muscle fibers, injected periurethrally as a potential treatment of stress urinary incontinence. We also studied myoblast injection into the bladder wall to potentially improve detrusor contractility. A myoblast cell line was transduced with adenovirus carrying the expression of the beta-galactosidase reporter gene while in culture. The cells were incubated with fluorescent latex microspheres (FLMs) to follow the outcome of the injected cells. The tissue was harvested 3-4 days after injection; sectioned, fixed, assayed for beta-galactosidase expression, and counterstained with H+E. Photographs of the slides were taken under light and fluorescence microscopy. We have noted a large number of cells expressing beta-galactosidase and containing FLMs in the urethral and bladder walls under fluorescent microscopy (8 animals). Many regenerative myofibers expressing beta-galactosidase were also seen in the urethral and bladder walls. The fusion of injected myoblasts to form myotubes was seen in both the urethral and bladder walls. The introduction of myoblasts into the urethral and bladder wall is feasible and results in formation of myotubes and myofibers in the smooth muscle layers of the lower urinary tract. We hypothesize that myoblast injections can be used as a non-allergenic agent to enhance urethral closure and bladder function.
Animals↗
Role of the forebrain in bladder overactivity following cerebral infarction in the rat.
This study was undertaken to investigate the contribution of the forebrain to bladder overactivity induced by cerebral infarction (CI). CI was induced by left middle cerebral artery (MCA) occlusion in female SD rat. Two and a half hours after CI or a sham operation (SO) decerebration was performed in some animals to eliminate forebrain influences on voiding function. Then bladder activity was monitored during continuous infusion cystometrograms in awake rats for 2.5 h. The effects of cumulative intravenous doses of MK-801 (0.1-1.4 mg/kg), an NMDA (N-methyl-D-aspartate) glutamatergic receptor antagonist, or sulpiride (0.1-41.1 mg/kg), D(2) selective dopaminergic receptor antagonists were studied over a 1.5-h period beginning 5 h after MCA occlusion. Bladder capacity was reduced by 57.5% after CI. In CI rats decerebration increased bladder capacity by 62.5% of predecerebration capacity. In SO rats bladder capacity was reduced by 25% after decerebration. MK-801 (0.4 and 1.4 mg/kg) increased bladder capacity in CI and CI-decerebrate rats, but did not change bladder capacity in SO-decerebrate rats. MK-801 decreased (60.7%) bladder capacity in SO-nondecerebrate rats. Sulpiride (11.1 and 41.1 mg/kg) significantly increased bladder capacity in CI, CI-decerebrate, and SO-decerebrate rats, but had no effect in SO-nondecerebrate rats. These results indicate that CI-induced decrease in bladder capacity is mediated by two mechanisms: (1) upregulation of an excitatory pathway from the forebrain, an effect blocked by decerebration and (2) downregulation of a tonic inhibitory pathway from the forebrain. The latter effect which can be induced by decerebration as well as CI unmasks a D(2) dopaminergic excitatory mechanism. An NMDA excitatory mechanism also contributes to the bladder overactivity after CI, but not after decerebration.
Animals↗